Figure 3. Inefficient activation of Cd8 genes in Brd1-deficient DP thymocytes.

(a) Cell cycle status of repopulated thymocytes. Percentages of cells in S phase in the indicated populations of thymocytes from the recipient mice in Fig. 2d were measured by detecting incorporated BrdU by flow cytometry at 2 h post-intraperitoneal injection of BrdU. Data are presented as mean±s.d. (Brd1 ^+/+, n =4; Brd1^Δ/Δ, n =3). (b) Hierarchical clustering of gene expression patterns of the indicated cell fractions obtained in microarray analyses. For clustering analysis, the Ward’s linkage rule was applied to all expressed genes in each of the sorted cells. The Ward’s linkage scores are indicated on the top. (c) A scatter diagram of microarray analysis. The average signal levels of indicated cell fractions compared with those of Brd1^Δ/Δ CD4 ⁺CD8 ⁻TCRβ^−/low cells are plotted. The upper and lower lines represent the borderline for a twofold increase and a twofold decrease, respectively. (d) Quantitative RT-PCR analysis of the expression of the genes indicated in Brd1 ^+/+ and Brd1^Δ/Δ thymocytes. Hprt1 was used to normalize the amount of input RNA. (e) Representative flow cytometric profiles of CD4 and CD8 expression in thymic T cells from 8- to 11-week-old Tie2-Cre (Brd1 ^+/+) and Tie2-Cre;Brd1^fl/fl (Brd1^Δ/Δ) mice expressing OT-I TCR transgene. Proportion of each gate is indicated. Data are representative of three independent experiments. (f) Representative flow cytometric profiles of CD4 and CD8 expression in PB of Brd1 ^+/+ OT-I and Brd1^Δ/Δ OT-I mice (left panels). Proportion of each gate is indicated. CD8 expression in PB T cells is also depicted in histograms (right panel).